Assessment of CD105, α-SMA and VEGF expression in gastric carcinomas.

In this study, we analyzed the microvessel density (MVD) for CD105+ and α-SMA+ vessels and the VEGF immunoexpression in 38 gastric carcinomas. CD105+ MVD had superior values at the advancing edge compared with the intratumoral area, no matter of the analyzed clinico-pathological parameters, the difference being significant only in intestinal type, moderate differentiated carcinomas as well as in T2-T3 carcinomas, without lymph node metastases (p<0.05). Intratumoral expression of CD105+ MVD indicated significant differences related to histological type (p=0.006), depth of invasion (p=0.027) and lymph node metastases (p=0.009), but without statistical association in case of the advancing edge or metastases. The assesses of α-SMA+ MVD indicated no differences between intratumoral and advancing edge areas, no matter of the analyzed parameters, excepting intestinal type carcinomas, which presented significant high values (p=0.003) at the advancing edge. VEGF score revealed significant differences related to histological type (p=0.020), differentiation degree of the intestinal type carcinomas (p=0.036) and depth of invasion (p=0.049). In case of metastases, the levels of VEGF expression were higher in the primary tumor, without statistically significant differences (p>0.05). It were significant differences of intratumoral VEGF expression depending on CD105+ MVD values (p=0.019), but not with α-SMA+ MVD (p>0.05). Angiogenesis evaluated through the VEGF and MVD (CD105+ and α-SMA+) expression is correlated with the progression and metastasis of gastric cancer and could be considered a prognostic marker of these tumors.

CD105 microvessels density, VEGF, EGFR-1 and c-erbB-2 and their prognostic correlation in different subtypes of cervical adenocarcinoma.

In this study, we have investigated the immunohistochemical expression of endoglin (CD105), vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR-1) and epidermal growth factor receptor 2 (c-erbB-2) and their prognostic correlation in 13 cases of cervical adenocarcinomas with mucinous, endometrioid and serous type differentiation. Our study revealed that for uterine cervix adenocarcinoma the most intense angiogenic activity occurs at the invasion front of these tumors. In addition, we noticed a trend towards increased CD105 MVD values in those cases in which were recorded the highest VEGF and c-erbB-2 reactivity. Thus, we concluded that in cervical adenocarcinomas occurs an intense process of angiogenesis, mainly at the invasion front, controlled by interrelations between VEGF and EGFR family members, especially the c-erbB-2 receptor. Further studies are needed to elucidate whether specific angiogenic molecular profiles exist in different histopathological subtypes of uterine adenocarcinomas and which is their impact on prognosis and therapeutic outcomes for these patients.

Immunohistochemical expression of EGF, c-erbB-2 and EGFR in intestinal variant of gastric adenocarcinomas.

In this study, we aimed to determine EGF, c-erbB-2 and EGFR expression in 25 specimens of intestinal gastric adenocarcinomas by standardized immunohistochemistry and to establish correlations with the major clinico-morphological parameters of these patients. We observed EGF reactivity in 22 (88%) cases, a c-erbB-2 protein expression in eight (32%) cases and an EGFR reactivity in 13 (46.42%) cases. The EGF expression was significantly correlated with the tumor degree of differentiation, but not with other investigated clinico-morphological parameters and nor with c-erbB-2 and EGFR1 expression. However, we noticed the existence of a dependence between c-erbB-2 and EGFR1 expression in the main tumor mass. Such immunoprofile suggests the possible intervention of autocrine and paracrine loops in the developing of intestinal variant of gastric adenocarcinomas.

Acinic cell carcinoma of the salivary glands: an immunohistochemical study of angiogenesis in 12 cases.

Acinic cell carcinoma (ACC) is the third most common epithelial malignancy of the salivary glands in adults, exhibiting a low-grade malignancy that mainly occurs in the parotid gland and at a relatively younger age than other salivary gland tumors. We performed an immunohistochemically study regarding angiogenesis in ACC, by assessing the CD105+ tumor microvessels density and investigating the VEGF and its receptors VEGFR1 and VEGFR2 expression in tumor samples. The results indicated an active angiogenesis in ACC, with the highest CD105-MVD score recorded in the solid variant. This fact was supported by the reactivity of tumor cells and endothelial blood vessel cells for VEGF and its receptors (VEGFR1 and VEGFR2). Thus, we concluded that in ACC do exist autocrine and paracrine VEGF loops implicated in growth and progression of this kind of salivary gland tumors.

Immunoexpression of Ki67 and cyclin D1 in oral squamous carcinomas.

In this study, we analyzed Ki67 and cyclin D1 immunoexpression in 44 oral squamous cell carcinomas from various anatomical sites. Ki67 immunoreaction was identified in all analyzed cases and presented an index of proliferation of 22% for well-differentiated carcinomas, 32% for moderately differentiated and 53% for the poorly differentiated ones. In case of cyclin D1, the mean positivity index was 8% for well-differentiated carcinomas, 18% for moderately differentiated and 34% for the poorly differentiated carcinomas. The analyzed biomarkers prove useful to identify lesions with poor differentiation and invasive behavior.

P53 and Ki67 immunoexpression in mucinous malignant ovarian tumors.

Ovarian cancer is the fourth cause of death by cancer among women and the first mortality cause in gynecological neoplasia. Our goal was to evaluate p53 and Ki67 immunoexpression and also the correlations with tumor stage and type. The study included 45 primary ovarian malignant mucinous tumors, diagnosed in patients in the IV and VI decade. From the standpoint of histopathology, there were 28 cases of borderline mucinous tumors and 17 mucinous carcinomas, predominantly stage I of the disease. The immunostaining for Ki67 was positive in all the cases, the highest levels being recorded in mucinous carcinomas (22.2% medium index) compared to the borderline tumors (9.5% medium index). Opposed to Ki67, the immunoreaction for p53 was present in 37.7% of all the tumors, predominantly in mucinous carcinomas where the stain has high values (52.3% medium index) in contrast with borderline lesions (15.5% medium index). The study indicated significant differences in p53 and Ki67 immunostain in relation to the tumor stage and histological type, there being a direct correlation of the expression of both proteins, in the studied tumors. P53 and Ki67 are useful markers for evaluating aggressive tumoral behavior and differentiating between mucinous carcinomas and borderline mucinous tumors.

The immunoexpression of EGFR and Her2/neu in oral squamous carcinoma.

In this study, we have analyzed the EGFR and Her2/neu expression in oral squamous carcinoma and adjacent dysplastic areas. The lesions were diagnosed especially in the sixth decade of life, in male patients, localized on the lips, especially as well and poorly differentiated carcinomas (34%). The EGFR immunostain has been intense in over 50% of the tumors cells in well-differentiated carcinomas, expression diminished in the moderately and poorly differentiated carcinomas. Her2/neu marker recorded a score of 3+ in moderately and poorly differentiated carcinomas, the reaction turning out positive in 25% of the cases.

Cervical adenocarcinoma: a retrospective clinicopathologic study of 16 cases.

Endocervical adenocarcinomas account for about 10-30% of cancers of the uterine cervix and display a variety of disparate morphologies. As an objective of the present work, we analyzed the clinicopathologic characteristics and prognostic factor of cervical adenocarcinoma. Clinicomorphological data of 16 cases of endocervical adenocarcinoma were reviewed during 2006-2011. Histopathologically, seven cases were of mucinous endocervical type, one intestinal type, two mucinous villoglandular type, four endometrioid type and two of serous type. The immunohistochemical investigation showed a tipically endocervical carcinoma profile ER-/PR-/Vim-/CEA+ in 10 cases (62.5%), which morphologically corresponded to: five mucinous endocervical type, one villoglandular type, three endometrioid type and one serous type. Regarding the prognosis we established that endometrioid endocervical adenocarcinoma is the histological variant with the worst prognosis, most cases been diagnosed in advanced stages (IIIA and IIIB) while at the opposite pole were papillary villoglandular and serous endocervical adenocarcinomas, diagnosed in less advanced stages of disease (IB and IIB). We concluded that the clinicomorphological diagnosis of endocervical adenocarcinoma is a challenging task, given to its multitude of histological variants and to the fact that immunohistochemistry investigations proved to be useful in only 63% of cases. In addition, we confirmed that the clinical stage is the most important prognostic factor and to some extent, the histomorphologic features can condition the biological behavior of these tumors.

Acinic cell carcinoma of the salivary glands: a retrospective clinicopathologic study of 12 cases.

Acinic cell carcinoma (ACC) is the third most common epithelial malignancy of the salivary glands in adults, with a low-grade malignancy that mainly occurs in the parotid gland and at a relatively younger age than other salivary gland tumors. We made a retrospective study on our acinic cell carcinoma casuistry aiming their clinico-pathological characterization and comparison with literature data. From 2000 through 2011 in our hospital were diagnosed only 12 cases of ACC. The clinico-epidemiological study revealed prevalence of these tumors in women, in the fourth decade of life and especially occurring in the parotid gland. The most common morphologic pattern of these tumors was a mixture of two or more variants with the solid/lobular and microcystic patterns more frequent associated. In 75% of investigated cases, the pTNM stage was I/II, with no cases of perineural or vascular invasion, but with lymph node dissemination presented in only three cases. Summing all these clinicopathological features, we conclude that for our casuistry the biological behavior of these tumors has been of low-grade malignancy.

IL-17 patterns in synovium, serum and synovial fluid from treatment-naïve, early rheumatoid arthritis patients.

BACKGROUND: There are actually becoming controversial data regarding the profiles of interleukin-17 (IL-17) in different pathogenical stages of rheumatoid arthritis (RA). OBJECTIVES: To assessing the IL-17 patterns in synovium, serum and synovial fluid from treatment-naïve early RA patients and to identifying potential correlations with disease activity markers and with synovial histopathological profile. MATERIALS AND METHODS: Serum samples from 30 treatment-naïve early RA patients were evaluated for C-reactive protein (CRP), erythrocytes sedimentation rate (ESR), rheumatoid factor (RF), anti-cyclic citrullinated peptide antibodies (anti-CCP). IL-17A levels were also assessed in serum and synovial fluid (SF). Disease activity score (DAS28) calculation was done for all patients. Control serum and SF samples were obtained from 29 patients with osteoarthritis (OA); control synovium specimens were obtained from eight patients with OA and during surgery for knee tear ligaments. Histopathological (Hp) score, immunohistochemical reactivity for IL-17 were also assessed in synovium of early RA patients and controls. Dependencies between serum and synovial profile of IL-17A and the other parameters were statistically tested. RESULTS: In early RA patients, strong correlations of serum and SF IL-17A levels were found with ESR, CRP, RF, anti-CCP, Hp score and IL-17 synovial immunoreactivity; a good correlation was noted with DAS28 score. Also, strong correlation was noted between serum and SF IL-17A levels. CONCLUSIONS: In early stages of untreated RA, simultaneous IL-17 assessment of serum, SF and synovium might be valuable in defining activity and predictive patterns, given that synovium is highly suggestive for an disease aggressivity and might express specific therapeutically targets.

P53, p16 and Ki67 immunoexpression in oral squamous carcinomas.

In this study, we have analyzed clinically, histopathologically and immunohistochemically a total of 34 cases of oral squamous carcinoma in 11 of the cases being identified adjacent epithelial dysplastic lesions. Carcinomas were diagnosed in patients aged 40-60 years, males, with chronic exposure to tobacco and/or alcohol, being located especially on the lips. Well-differentiated carcinomas have been predominant (52.9%) in stage I/II tumoral (88.3%). Immunoexpression analysis of p53, p16 and Ki67 did not reveal statistically significant differences between the expression of markers and clinical or histopathological parameters, except Ki67 whose increased expression was associated to the decrease of the degree of tumoral differentiation and with high degree dysplasia. The positivity index and the intensity of reaction were increased at the level of dysplasic epithelium for p16 and at the level of tumoral invasion front for the p53 and Ki67. The study highlights the value of the immunostain for p16 in identifying dysplasic lesions and predictive importance of p53 and Ki67 markers in identifying the aggressive forms of oral carcinomas.

Expression of Epidermal Growth Factor (EGF) and its receptors (EGFR1 and EGFR2) in chronic bronchitis.

Chronic Obstructive Pulmonary Disease (COPD) is one of the leading causes of morbidity and mortality in the industrialized and in the developing countries. It is believe, at least in part, that some of the structural changes that occur in COPD would be a result of epidermal growth factors (EGFs) and their receptors. Therefore, our study aims was to examine the expression patterns of EGF and their receptors (EGFR1 and c-erbB2) in the bronchial mucosa from the biopsy specimens harvested from smoking and non-smoking CB patients, compared with their expression in normal controls. The statistical analysis proved that for both EGF and EGFR1 reactivity were significant correlation with smoking status and FEV1% scores. Thus, we found that the highest levels of its expression were recorded in smoker CB patients with higher FEV1% scores. Regarding cellular localization and staining pattern, we noticed a cytoplasmic and nuclear immunostaining for EGF in bronchial epithelium both for control and CB subjects at the level of basal and ciliated cells. For the receptors, reactions were at the membrane level especially at the lower lateral junctions between ciliated cells and their junctions with basal cells. This reactivity proved the pathogenic implication of the EGF and their receptors in patients with CB and suggests that blockade of the EGFR pathway can be an alternative successful therapy.

Immunohistochemical expression of MMP-9, TIMP-2, E-cadherin and vimentin in ameloblastomas and their implication in the local aggressive behavior of these tumors.

Among the benign epithelial odontogenic tumors, the ameloblastoma occupies a special place because of their local aggressive behavior with devastating jaw destructions, which lead to a high recurrence rate, even following the radical surgery. In an attempt to clarify the mechanisms underlying this behavior we immunohistochemically investigated the reactivity of different histological variant of ameloblastoma to MMP-9, TIMP-2, E-cadherin and vimentin. A semiquantitative assessment of their reactivity in the epithelial neoplastic compartment was done and statistical correlation was attempt with histological variant and between them. The MMP-9 and TIMP-2 reactivity was observed with variable intensity both in the neoplastic epithelium and in the stroma that surround the ameloblastic proliferations. Only for MMP-9, the statistical tests proved the existence of significant differences within major ameloblastic histological variants, with the highest reactivity acanthomatous type. The stroma independent of the histological variant had the highest reactivity at the invasive front adjacent to the tumoral islands. Immunoreactivity for E-cadherin was more obvious in the follicular type at the level of stellate-reticulum like cells, and decreased in the peripheral columnar cells, as they are closer to the invasion front. Vimentin reactivity was present in the neoplastic epithelium only in the peripheral columnar cells at the invasion front but at this site, the stroma had the highest expression. Statistical analysis proved the existence of inverse relationship between MMP-9 and E-cadherin scores, while vimentin score had an inverse relationship both with TIMP-2 and E-cadherin scores. Our results proved the implication of these four markers in the growth of ameloblastomas and as a consequence their utility in monitoring of local aggressiveness of these tumors.

VEGF and his R1 and R2 receptors expression in mast cells of oral squamous cells carcinomas and their involvement in tumoral angiogenesis.

More than 90% of malignant neoplasms of the oral cavity are squamous carcinomas of oral mucosa and most are conventional type. This study included 60 cases of oral squamous carcinoma processed by usual histological technique and stained with Hematoxylin-Eosin and Alcian Blue-Safranin (Alcian Blue powder from Sigma Aldrich, code A5268-25G) and diagnosed in the Department of Pathology, University of Medicine and Pharmacy of Craiova. Double immunohistochemical (CD105-tryptase, VEGF-, VEGFR1-, VEGFR2-tryptase) or combined histochemical and immunohistochemical reactions (VEGF÷ VEGFR1÷ VEGFR2÷Alcian Blue-Safranin) followed the particular morphological aspects of the mast cells, their relation with the blood vessels and the overlap signal of tryptase and Alcian Blue-Safranin÷VEGF and its R1 and R2 receptors in mast cells. Immunostaining for VEGF and its R1 and R2 receptors was present both in tumor cells and mast cells. Double immunohistochemical-histochemical reactions allowed us angiogenic profiling the mast cells. The signal overlap was present for VEGF-, VEGFR1-, VEGFR2-tryptase intratumoral and tumor invasion front mast cells. Student's t-test for comparison of intratumoral and the invasion front MDM showed highly significant value (p=3.23 E-08). VEGF÷ VEGFR1÷ VEGFR2÷Alcian Blue-Safranin revealed particular morphological aspects of mast cells, in with different morphology, shapes, sizes and degrees of degranulation. Statistical analysis showed a linear correlation between MDM and MVD inside the tumor (Pearson coefficient =0.47) and a weak linear correlation at the front of invasion (Pearson coefficient =0.19). This study has highlighted the importance of mast cells in the tumor growth of the oral squamous carcinomas, especially in terms of their proangiogenic profile (expression of VEGF and its R1 and R2 receptors). In addition, their quantification as MDM makes this parameter a useful prognostic marker.

VEGF expression in human brain tissue after acute ischemic stroke.

Ischemic stroke is the third most common cause of death in humans, requiring further studies to elucidate its pathophysiological background. One potential mechanism to increase oxygen delivery to the affected tissue is induction of angiogenesis. The most potent proangiogenic factor is VEGF. For this reason, our study investigated immunohistochemically VEGF reactivity in different cellular brain compartments from 15 ischemic stroke patients, as well as from 2 age control cases. By enzymatic immunohistochemistry, we investigate VEGF expression in different brain cell compartments and then we quantified its signal intensity by assessing integrated optical densities (IOD). To establish the exact cellular brain topography of VEGF immunoreactivity we performed double fluorescent immunohistochemistry series (VEGF÷NeuN, GFAP, CD68, CD105). In control samples, VEGF reactivity was observed especially in neurons from the Brodmann cortical layers IV to VI and in protoplasmic astrocytes from the deeper layers of gray matter and in endothelial cells from normal blood vessels because of systemic hypoxia generated after death. In acute ischemic stroke samples, this reactivity was noticed in all brain cellular compartments but with different intensities. The most reactive compartment was the neurons, the intensity of VEGF reaction decreasing with the lesional age from the core infarct toward intact adjacent brain cortex. With a lower intensity, VEGF reaction was noticed in astrocytes compartments, especially in gemistocytic astrocytes adjacent to the liquefaction zone. We also noticed a weak reaction in activated non-phagocytic microglia from the periphery of liquefaction zones, and high VEGF-CD105 colocalization values at the level of microvessels that surround the infarcted brain area. In conclusion, this reactivity could suggest that VEGF might exhibit neuronal and glial protective effects and also a neoangiogenic property in acute ischemic stroke, facts that may have significant therapeutically impact on these patients.

The utility of CD44, CD117 and CD133 in identification of cancer stem cells (CSC) in oral squamous cell carcinomas (OSCC).

One of the theories regarding oral carcinogenesis is that the tumor growth is dependent on cancer stem cells (CSCs) that have the capacity of self-renewal and of giving rise to more differentiated tumor cells, like the stem cells do in normal tissues. The most used methods of CSCs isolation are based on their identification based on the expression of different cell surface markers. The markers qualified for this purpose have been described originally in studies involving hematopoietic or embryonic stem cells. Thus, we were interested to study the expression of the most used CSCs surface markers for formalin fixed paraffin embedded tissue samples from oral squamous cell carcinoma (OSCC). We investigated by immunohistochemistry thirty tissue samples of OSCCs with different degrees of differentiation and different oral locations. We were interested to establish the tissular localization pattern for cells expressing CD44, CD133 and CD117 in tumoral samples. The results indicated that with the exception of CD44, the other two surface markers were expressed only in tumoral stromal cells. When we looked at their origin (by double immunohistochemistry for cytokeratins AE1-AE3, vimentin and CD34) we concluded that they are of mesenchymal nature. Also, we proved that some of these cells also co-expressed CD44 but were negative for CK5÷6. Moreover, some of the stromal cells that were positive to CD133 and especially for CD117 also had reactivity to tryptase showing their mast cell nature. In conclusion, our study proved that CD44 has limited utility in identifying oral CSCs, while CD117 and CD133 expression appears to be limited more in identifying mesenchymal stem cells.

E-cadherin in gastric carcinomas related to histological prognostic parameters.

AIM: The immunohistochemical study of E-cadherin in gastric carcinomas, related to tumor aggressiveness factors (invasive and metastatic potential). MATERIALS AND METHODS: The studied material was gastric resection specimens taken from of 60 patients with gastric cancer, during 2009. The tissue was processed using standard histopathological technique, which allowed the assessment of the well-known morphological parameters of prognostic value. Later on the specimens has undergone immunohistochemical processing for E-cadherin (NCH-38 clone), to evaluate its expression in relation with these prognostic parameters. RESULTS: E-cadherin was positive in 65% from gastric carcinomas, with highest positivity index for well (80% cases) and moderate (17.64% cases) differentiated intestinal type tumors, while a large number of poorly differentiated tumors (55.55%) were E-cadherin negative. Among diffuse type carcinomas, the majority of advanced stage tumors (50% of serosal invasive tumors and 100% of peritoneal disseminated tumors) and also a high number of tumors with vascular and lymphatic invasion (50% and respective 80% cases) represented the E-cadherin negative category (54.54%). The E-cadherin staining was also negative in 75% of lymph node positive diffuse type carcinomas and in all metastatic tumors. CONCLUSIONS: We found that irrespective of histologic type, the E-cadherin expression was reduced to negativity in advanced stages of gastric carcinoma.

Uterine pseudotumors.

Pseudotumoral uterine lesions include benign reactive and artefactual changes, epithelial and mesenchymal, which occasionally are incorrectly interpreted as malignant or even premalignant lesions. Although some of these changes may have architectural or cytological abnormality, they are different from those observed in premalignant and malignant uterine lesions. The most common pseudotumoral lesions of the endometrium include various types of epithelial and stromal metaplasia, pseudolymphomas, inflammatory pseudotumor, adenomyosis, post-therapy surgical changes, artefactual changes, etc. Most of these changes may coexist with endometrial hyperplasia or endometrial carcinoma, and also with some benign conditions such as polyps or in combination with hormonal therapy or even in normal cyclic endometrium. These associated endometrial changes may raise important issues regarding the diagnosis and subsequent therapy.

Mucinous adenocarcinoma of the colon - a histochemical study.

Colorectal carcinoma is a major cause of cancer associated with a high rate of morbidity and mortality in the western world. One of the pathologic features considered to be important for prognostic is mucin production. Many authors confirmed that colon carcinomas with high mucin content tend to re-occur locally and carry a poor prognosis. For histochemical evaluation of mucin content, we investigated 149 patients who underwent surgical resection of sporadic colon adenocarcinomas, all over a 5-year period. For histological classification we used the WHO recommendation (2000) and to be more accurate we sub-classified mucinous adenocarcinomas by morphometrical analysis in three categories: pure mucinous, with extracellular mucin more than 80% of the tumoral volume; mixed type, with 50-80% extracellular mucin; and mixed type with less than 50% extracellular mucin. For histochemical investigation, we used stains such as: mucicarmine, PAS ÷ Alcian Blue and High Iron Diamine ÷ Alcian Blue. Our study proved the predominance of mixed mucinous adenocarcinomas with less than 50% extracellular mucin, followed by the pure mucinous type. From the biochemical composition's point of view, the predominant cases were those with acidic mucins, especially in pure mucinous adenocarcinomas (>90%), while those with mixtures of acidic and neutral mucins were present in 62% of the cases. In addition, our study showed the prevalence of sialomucins over sulphomucins (68%), particularly in pure mucinous adenocarcinomas (77%). Clinical pure mucinous forms were detected mainly in advanced stages, but in terms of lymph node metastasis rate, they were secondary after mixed type with 50-80% extracellular mucin.